柑桔链格孢褐斑病菌种群对嘧菌酯的抗性监测和抗性分子机制

Molecular Mechanism and Monitoring the Azoxystrobin Resistance in Citrus Alternaria alternata Population in China

为明确抗嘧菌酯（azoxystrobin）柑桔链格孢褐斑病菌在我国的分布、频率、抗性水平和抗性分子机制,从重庆、湖南、浙江、云南、广东和广西等有链格孢褐斑病发生的柑桔产区,收集了54个链格孢褐斑病菌（Alternaria alternata）菌株,通过刃天青染料显色法测定各菌株孢子萌发和幼菌丝生长对嘧菌酯的敏感性,扩增、测序和比较嘧菌酯敏感和抗性菌株嘧菌酯靶标Cytb基因部分序列。结果表明,嘧菌酯抗性菌系在各采样产区均存在,抗性频率在14%~42%之间,平均为30%,抗性系数为18~112。2个敏感菌株的Cytb基因编码的第143位氨基酸为甘氨酸,16个抗性菌株则为丙氨酸。因此认为,抗嘧菌酯的链格孢褐斑病菌在有褐斑病发生的地区普遍存在,抗性分子机制是Cytb基因的第143位氨基酸从甘氨酸突变为丙氨酸。基于本研究结果,建议在防治柑桔链格孢褐斑病时需谨慎选择QoIs类杀菌剂。

To understand the distribution,frequency,level and the molecular mechanism of azoxystrobin-resistance of Alternaria alternata,the causal agent of citrus alternariabrown spot（ABS）in China,54 A.alternata strains were collected from citrus orchards in Chongqing,Hunan,Zhejiang,Yunnan,Guangdong and Guangxi provinces,where ABS outbreaks.The sensitivities of each strain to azoxystrobin were tested using resazurin-based microtiter assay.The molecular mechanism of azoxystrobin-resistance was determined by comparison of amplified partial sequences of azoxystrobin target gene Cytbfrom 2azoxystrobin-sensitive and 16azoxystrobin-resistant strains.The results showed that the azoxystrobin-resistant strain is commonly present in all those provinces.The frequency of resistant strains ranged from 14%to 42%,with an average at 30%,and the resistant coefficient ranged between 18 and 112.Comparing the protein sequences of Cytb gene among 2sensitive and 16 resistant strains revealed a single mutation occurred at position 143,which is glycine in azoxystrobin-sensitive strains,but alanine in resistant strains,suggesting the substitution of glycine to alanine at 143 of Cytb could confer A.alternata with azoxystrobin resistance.This result is in accordance to the previous reports.The common presence of azoxystrobin-resistant A.alternata strain in China reminded us that to control ABS,we need to be cautious on selection of QoI fungicides.